Homologous recombination(HR)is one of major mechanisms that repair DNA double strand breaks(DSBs)and helps maintain genome stability.Many genes involved in HR contribute to hereditary and spontaneous cancer risk when deficient.In particular,germline mutations of the HR genes BRCA1 and BRCA2 account for 5-10%of familial breast cancers and 10-18%of familial ovarian cancers.HR defects caused by deficiency of BRCA1/2 and many other HR genes render cells hypersensitive to PARP inhibitors(PARPi),leading to the development of PARPi as effective “synthetic lethal” therapeutics in treatment of HR-deficient cancers.However,many patients with HR defects are not as sensitive to PARPi as expected.We speculate that these patients may maintain a certain level of HR activity that can offset the efficacy of PARPi.In fact,using a novel and quantitative HR reporter system,we have previously shown: 1)H2AX is required for efficient HR and phosphorylation of S139 on its SQEY motif is essential for this HR function;2)H2AX-dependent HR requires the interaction of pSQEY of phosphorylated H2AX(also termed “γH2AX”)with tandem BRCT domain of MDC1;3)H2AX/MDC1-mediated HR remains active in BRCA1-and ATM-deficient mouse embryonic stem(ES)cells;and 4)Loss of H2AX-dependent HR enhances hypersensitivity of ATM-deficient ES cells to PARPi.These findings suggest that inhibiting H2AX/MDC1-mediated HR by blocking the γH2AX-MDC1 interaction may improve the efficacy of PARPi-based cancer therapy.We thus used computer-aided drug design to target the γH2AX-interacting BRCT domain of MDC1 and perform virtual ligand screening of ChemDiv chemical library(about 1,500,000 small molecules)for γH2AX-MDC1 interaction inhibitors.Among 132 small molecule compounds selected,we tested 80 for their effects on HR and found 4 compounds with an inhibitory effect on HR.We are currently determining whether these 4 compounds indeed target the γH2AX-MDC1 interaction,thus inhibiting H2AX/MDC1-mediated HR.We hope to identify 1-2 lead compounds that can be developed into effective therapeutics for treating cancer patients with HR-deficient tumors in combination with PARPi.